Virus-infected macrophages are the major therapeutic target in the brain tissue, even though the productive virus replication in these cells is necessary but not sufficient to cause HIV Associated Dementia (HAD). HAART is still generally recommended for patients with AIDS and in general this treatment has been effective in decreasing the incidence of HAD; however a significant group of patients remains refractory to HAART even when its CNS penetration is enhanced by supplementing drugs. For this reason alone, the activation of native cellular mechanisms inducing anti-viral resistance in cells migrating to various organs including CNS should be considered a viable alternative to add to the existing antiviral approaches for HAD. We have induced such responses mediated through soluble molecule [previously named HRF (HIV-1 Resistance Factor)] able to prevent virus replication in CD4+ T cells. Recently we identified the HRF as a truncated form of a potent transcription suppressor - CTCF (tCTCF). The biological activity of tCTCF was confirmed by several experimental methods: (1) the siRNA silencing of CTCF mRNA in HRF(+) cells reversed the HIV-1 resistance phenotype and reduced the biological activity of their cell culture supernatants; (2) depletion of tCTCF from HRF(+) cell culture supernatant abolished its antiviral activity; while (3) the affinity purified tCTCF induced antiviral activity in human macrophages; and (4) the affinity purified tCTCF inhibited the transcription of HIV-1 LTR promoted expression of reporter gene. Our preliminary data showed that upon HIV-1 exposure, CTCF from nuclear extracts of HRF(+) but not HRF(-) control cells bound to DNA probe comprising NF-B, Sp1 and YB-1 binding sequences, possibly through its interaction with YB-1 protein. We hypothesize that since NF-B and YB-1 binding sites are separated by only 6 nucleotides the CTCF/YB1 complex bound to the virus promoter might obstruct the formation of NF- B/DNA binding, thus halting transcription of virus genome. Taken together we discovered a novel form of CTCF protein that mediates innate responses to HIV-1 in human macrophages. The main goal of this application is to prepare foundation for future delivery of tCTCF as a biotherapeutic for treatment of HIV-1 in CNS. In this grant application we propose: (1) To study the mechanism of tCTCF mediated cellular responses to HIV-1; (2) To determine the minimum tCTCF amino-acid sequence required for the induction of antiviral responses and efficacy assessment of tCTCF treatment in primary human macrophages; and (3) To determine the minimal CTCF toxicity and brain entry in vivo.